Mouse and human iPS cells were established recently. Takahashi and Yamanaka (Non patent literature 1) established mouse iPS cells by introducing Oct3/4, Sox2, Klf4 and c-Myc genes into fibroblasts derived from a reporter mouse in which a neomycin-resistant gene was knocked-in into the Fbx15 locus, and forcibly expressing the four genes. Okita et al. (Non-patent literature 2) prepared a transgenic mouse by integrating green fluorescent protein (GFP) and puromycin-resistant genes into the Nanog locus, forcibly expressed the above four genes in the fibroblasts derived from the transgenic mouse, and selected puromycin-resistant and GFP-positive cells, thereby successfully generated iPS cells (Nanog iPS cells) which are almost equal to embryonic stem (ES) cells in gene expression and epigenetic modification. Nanog is specifically expressed in pluripotent cells and is more limitedly expressed in pluripotent cells than Fbx15. Similar results were also reproduced by other study groups (Non-patent literatures 3 and 4). Thereafter, it was found that iPS cells can be prepared even using three genes of Oct3/4, Sox2 and Klf4, i.e. the above four genes except for c-Myc gene (Non-patent literature 5).
Moreover, Takahashi et al. (Non-patent literature 6) successfully generated human iPS cells by introducing the four genes similar to those used for generating mouse iPS cells into fibroblasts derived from human skin. On the other hand, Yu et al. (Non-patent literature 7) prepared human iPS cells using Nanog and Lin28 instead of Klf4 and c-Myc. Furthermore, Park et al. (Non-patent literature 8) prepared human iPS cells by using TERT known as a human cell-immortalizing gene and SV40 large T antigen, in addition to the four genes of Oct3/4, Sox2, Klf4, and c-Myc. As described above, it was shown that iPS cells which are comparable to ES cells in pluripotent differentiation can be prepared in human and mouse by introducing particular factors into somatic cells.
The c-Myc gene has the risk of tumor development, and therefore, no use of the gene is desirable in the induction of iPS cells that are used for clinical therapies. Since, however, iPS cell generation efficiency was reported very low when using the three genes without c-Myc (Non-patent literature 5), it has been thought that research of a factor to replace the c-Myc is important. The factor hopefully has an effect to improve iPS cell generation efficiency comparable to or greater than that of c-Myc, and has a reduced risk of tumor development.
In this connection, the present inventors have previously revealed that iPS cell generation efficiency was improved by using L-Myc instead of c-Myc in the generation of human iPS cells. In addition to iPS cell generation efficiency, L-Myc was revealed to increase surviving days of chimeric mice and decrease tumor formation (patent literatures 1 and 2).
Documents cited (the following documents are herein incorporated by reference):
Non patent literature 1. Takahashi, K. and Yamanaka, S., Cell, 126: 663-676 (2006)
Non patent literature 2. Okita, K. et al., Nature, 448: 313-317 (2007)
Non patent literature 3. Wernig, M. et al., Nature, 448: 318-324 (2007)
Non patent literature 4. Maherali, N. et al., Cell Stem Cell, 1: 55-70 (2007)
Non patent literature 5. Nakagawa, M. et al., Nat. Biotethnol., 26: 101-106 (2008)
Non patent literature 6. Takahashi, K. et al., Cell, 131: 861-872 (2007)
Non patent literature 7. Yu, J. et al., Science, 318: 1917-1920 (2007)
Non patent literature 8. Park, I. H. et al., Nature, 451: 141-146 (2008)
Patent literature 1. U.S. Publication No. 2009-0227032
Patent literature 2. International Publication No. WO2009/057831